The invention relates to macrolides, particularly to macrolactam macrolides. It concerns the compounds of formula I 
wherein either
R1 is hydroxy in the xcex2-configuration; and
R2 is methyl and
one of R3 and R4 is ethyl and the other is methyl; or
R1 is chloro in the xcex1-configuration; and
one of R2, R3 and R4 is ethyl and the others are methyl;
in free form and, where such forms exist, in salt form, hereinafter briefly named xe2x80x9cthe compound of the inventionxe2x80x9d.
When R1 in formula I is in the xcex1-configuration it is above, and when it is in the xcex2-configuration it is below, the plane of the paper. R1 preferably is chloro in the xcex1-configuration. R3 or R4, particularly R3, preferably is ethyl.
The compounds of the invention are hereinafter briefly named as follows:
when R1 is hydroxy and
R3 is ethyl: 21-ethyl-ascomycin; or
R4 is ethyl: 27-ethyl-ascomycin;
when R1 is chloro and
R2 is ethyl: 19-ethyl-ASM; or
R3 is ethyl: 21-ethyl-ASM; or
R4 is ethyl: 27-ethyl-ASM;
i.e. they are, respectively:
(1R,9S,12S,13R,14S,17R,18E,21S,23S,24R,25S,27R)-12-[(1E-2-[(1R,3R,4R)-4hydroxy-3-methoxycyclohexyl]-1-methylvinyl]-17,21-diethyl-1,14-dihydroxy-23,25-dimethoxy-13,19,27-trimethyl-11,28-dioxa-4-azatricyclo-[22.3.1.04,9]octacos-18-ene-2,3,10,16-tetrone;
(1R,9S,12S,13R,14S,17R,18E,21S,23S,24R,25S,27R)-12-[(1E-2-[(1R,3R,4)-4-hydroxy-3-methoxycyclohexyl]-1-methylvinyl]-17,27-diethyl-1,14-dihydroxy-23,25-dimethoxy-13,19,21-trimethyl-11,28-dioxa-4-azatricyclo-[22.3.1.04,9]octacos-18-ene-2,3,10,16-tetrone;
(1R,9S,12S,13R,14S,17R,18E,21S,23S,24R,25S,27R)-12-[(1E-2-[(1R,3R,4S)-4-chloro-3-methoxycyclohexyl]-1-methylvinyl]-17,19-diethyl-1,14-dihydroxy-23,25-dimethoxy-13,21,27-trimethyl-11,28-dioxa-4-azatricyclo-[22.3.1.04,9]octacos-18-ene-2,3,10,16-tetrone;
(1R,9S,12S,13R,14S,17R,18E,21S,23S,24R,25S,27R)-12-[(1E-2-[(1R,3R,4S)-4-chloro-3-methoxycyclohexyl]-1-methylvinyl]-17,21-diethyl-1,14-dihydroxy-23,25-dimethoxy-13,19,27-trimethyl-11,28-dioxa-4-azatricyclo-[22.3.1.04,9]octacos-18-ene-2,3,10,16-tetrone; and
(1R,9S,12S,13R,14S,17R,18E,21S,23S,24R,25S,27R)-12-[(1E-2-[(1R,3R,4S)-4-chloro-3-methoxycyclohexyl]-1-methylvinyl]-17,27-diethyl-1,14-dihydroxy-23,25-dimethoxy-13,19,21-trimethyl-11,28-dioxa-4-azatricyclo-[22.3.1.04,9]octacos-18-ene-2,3,10,16-tetrone.
21-Ethyl-ascomycin and 27-ethyl-ascomycin are higher homologues of ascomycin. 19-Ethyl-ASM, 21-ethyl-ASM and 27-ethyl-ASM are higher homologues of pimecrolimus (ASM) (33-epichloro-33-desoxy-ascomycin).
The compounds of the invention can be prepared by a process comprising
a) for the preparation of the compounds of formula I as defined above wherein R1 is hydroxy in the xcex2-configuration,
xe2x80x83cultivating an appropriate microorganism and isolating from the resultant culture medium the corresponding compounds of formula I wherein R1 is hydroxy in the xcex2-configuration; or
b) for the preparation of the compounds of formula I as defined above wherein R1 is chloro in the xcex1-configuration,
xe2x80x83replacing under epimerization hydroxy with chloro in a corresponding compound of formula I wherein R1 is hydroxy in the xcex2-configuration,
and recovering the resultant compounds in free form or, where such forms exist, in salt form
The process of the invention is effected in conventional manner.
In variant a) any ascomycin-producing microorganism strain may be used which produces higher homologues of ascomycin, e.g. as impurities, preferably a Streptomyces hygroscopicus strain. Such strains are known and available from public depositories. Preferably, strains are used which produce significant amounts of higher homologues of ascomycin, or cultivation conditions are chosen which allow preparation of enhanced amounts of higher homologues of ascomycin. Such strains are known and readily accessible, such as Streptomyces hygroscopicus subsp. ascomyceticus (e.g. ATCC 14891, ATCC 53855, ATCC 55087, ATCC 55276, ATCC 55558, DSM 5085), Streptomyces tsukubaensis No. 9993 (FERM BP-927), Streptomyces hygroscopicus subsp. yakushimaensis No. 7238 (FERM BP-928, NRRL 18488); or natural or artificial mutants thereof, and cultivation conditions for enhanced higher homologues production are known or can readily be determined in conventional manner therefrom Conveniently, appropriate mutant strains may be created exhibiting enhanced higher homologues production, or selected, in conventional manner, or culture may be effected under modified conditions, such as with increased concentration of the C4 precursor sodium butyrate in the culture medium.
Variant b) is a substitution reaction under simultaneous or concomittant epimerization. It is effected e.g. as described in EP 427680. It preferably is effected in an inert solvent such as tetrahydrofurane or toluene. Preferably the reaction is effected with tetrachloromethane or N-chloro-succmimide in the presence of triphenylphosphine, conveniently in an alkaline medium such as collidine.
The resultant compounds of the invention may be isolated from the cultivation or reaction mixture and purified in accordance with known methods. However, it has been found that, surprisingly, use of a chiral stationary phase such as Kromasil(copyright) during chromatographic purification may greatly facilitate isolation of, in particular, the compounds of the invention wherein R1 is chloro.
The starting materials and intermediate compounds are either known or can be prepared according to known methods or analogously to known methods or analogously as described in the Examples.